Sample Preparation for Global Metabolic Profiling of Vaccinia Virus-Infected Primary Human Foreskin Fibroblasts.

Vaccinia virus (VACV), the prototype member of the Poxviridae family, has played a crucial role in medicine as a key component in the development of smallpox vaccines, contributing to the eradication of this deadly disease. Beyond its historical significance, VACV continues to be pivotal in researching metabolic alterations induced by viral infections. Studies have revealed that VACV can impact pathways such as glycolysis, the tricarboxylic acid (TCA) cycle, and lipid metabolism in host cells, offering valuable insights into host-virus interactions and broader cellular metabolism. The preference for primary cells, such as human foreskin fibroblasts (HFFs), over cancer cells in metabolic studies is justified for their physiological relevance, representing native cell types with genetic stability. Metabolic profiling is an ideal tool for studying virus-induced metabolic alterations, providing a comprehensive analysis of changes in cellular metabolism triggered by viral infections. This chapter outlines a protocol for extracting HFFs, culturing, infecting them with VACV, and conducting untargeted global metabolic profiling to elucidate detailed metabolic statuses of the infected cells. This protocol may be modified for understanding the intricacies of host-virus interactions at the metabolic interface for other poxviruses and non-poxviruses.